Numerous classes of bacteria and virus particles include proteins that undergo transitions from soluble to membrane inserted forms, which can severely compromise a human cell and the human organism in general. Often, the change in protein conformation from water soluble to membrane inserted (e.g., membrane soluble) occurs by the protein opening to expose hydrophobic motifs (e.g., amino acids, peptides, or polypeptides) that are usually within the protein core. The change in protein conformation from water soluble to membrane inserted configurations can occur during acidification, such as occurs in an endosome or lysosome. As such, inhibiting the change in protein conformation from water soluble to membrane inserted may be useful for inhibiting the negative consequences, such as disease or compromised health, which may arise from such a change in protein conformation.
Currently, some high-throughput technologies that are used to identify small molecule protein stabilizers rely on processes that heat and denature the target protein that may change conformation. Unfortunately, heating and denaturing proteins may be prone to artifacts ranging from heat induced aggregation and non-equilibrium transitions (i.e., dye influence equilibrium). Also, such heating and denaturing techniques are not suitable for many proteins. One of the most problematic issues with current techniques includes the potential ligands being identified under non-physiological temperatures. No high throughput systems exist where the screens to identify protein stabilizers are performed under physiological or near physiological conditions. Furthermore, current techniques include non-equilibrium processes and destroy the target protein. The identified ligands must bind to the protein under non-physiological conditions and these physical constraints often results in false positive and negative results.
One approach to inhibit such protein conformation transformations can include preventing an acid dependent transition, which may be accomplished using general or more specific protein stabilizers. Therefore, there is a need in the art for methods for identifying and testing protein stabilizers that can inhibit protein conformation transformations, such as those that occur with bacterial toxins and viral particle proteins.